As a result of progress made in the field of gene recombination technology, it has become possible to introduce a gene encoding a target protein into cultured cells or cells in a plant body and express the protein in those cells. In general, when expressing protein by introducing a gene into the nuclear genome of a plant, in the case of having only introduced the genetic region that encodes the target protein, the expressed protein accumulates in the cytoplasm. In addition, by introducing a gene that encodes a protein in which several amino acids referred to as a localization signal have been added to the N-terminal and C-terminal of the target protein, the expressed protein can be made to migrate to and accumulate in locations referred to as intracellular organelles, such as endoplasmic reticulum (ER), vacuoles or chloroplasts, or in an extracellular region (apoplast) (see Non-Patent Document 1). In addition, a technology is also known that consists of introducing a gene that encodes a target protein directly into a chloroplast genome, and allowing the entire process from gene expression to protein accumulation to take place within the chloroplast (see Non-Patent Document 2).
In particular, vacuoles are intracellular organelles that have the largest volume among plant cells, and are expected to serve as favorable locations for accumulation of foreign protein. A target protein can be localized in a vacuole by adding an amino acid sequence referred to as a vacuole localization signal to the N-terminal or C-terminal of the target protein. In actuality, proteins required during germination are specifically accumulated in vacuoles in seeds, and these vacuoles are referred to as protein storage vacuoles. For example, a transgenic plant is known in which foreign cellulase is accumulated in the storage vacuoles of seeds (see, for example, Non-Patent Document 3). However, since seeds only occupy a small proportion of plant biomass, there is the problem of the amount of protein being small relative to the total amount of biomass produced.
In order to produce a large amount of protein in a plant body, it is desirable to accumulate protein in vacuoles occupying a large volume in vegetative organs accounting for the majority of plant biomass, and particularly in tissues such as leaves or stems located above ground that can be harvested easily. However, proteolytic vacuoles rich in proteases are present in the cells of vegetative organs such as leaves or stems. Consequently, these vacuoles are not suitable for protein accumulation, and even if a foreign protein was localized in the vacuoles of leaves or stems using a vacuole localization signal, it would be extremely difficult to accumulate that foreign protein at high concentrations (see, for example, Non-Patent Documents 4 and 5).
On the other hand, myrosin cells are a type of atypical cell having a unique form present in plants, and these myrosin cells are present in leaves and are capable of accumulating large amounts of protein in their vacuoles. The myrosin cells present in the leaves of plants of the order Capparales, which includes the Cruciferae family, are known to specifically accumulate the vacuoles thereof a protease thought to be involved in the plant's defense against disease and harmful insects in the form of thioglucoside glucohydrolase (TGG, also known as myrosinase) (see, for example, Non-Patent Document 6). In wild thale cress plants (Arabidopsis thaliana) belonging to the order Capparales, Brasiccaceae family, although myrosin cells are only present in limited regions surrounding vascular bundles, Arabidopsis thaliana mutant vam3-4/ssm and Arabidopsis thaliana mutant vam3-3 have been found that have increased numbers of myrosin cells and contain accompanying increased amounts of myrosinase in those cells (see, for example, Non-Patent Documents 7 and 8). Mutant atvam3-4/ssm is a mutant in which a frame shift has occurred due to the absence of 34 bp in the 6th intron of vam3 gene, while mutant atvam3-3 is a mutant deficient in the vam3 gene. In addition, Arabidopsis thaliana mutants tgg1 and tgg2 are also known that are deficient in myrosinase present in the vacuoles of myrosin cells (see, for example, Non-Patent Document 9).